RESUMO
Tobacco misuse as a comorbidity of schizophrenia is frequently established during adolescence. However, comorbidity markers are still missing. Here, the method of label-free proteomics was used to identify deregulated proteins in the medial prefrontal cortex (prelimbic and infralimbic) of male and female mice modelled to schizophrenia with a history of nicotine exposure during adolescence. Phencyclidine (PCP), used to model schizophrenia (SCHZ), was combined with an established model of nicotine minipump infusions (NIC). The combined insults led to worse outcomes than each insult separately when considering the absolute number of deregulated proteins and that of exclusively deregulated ones. Partially shared Reactome pathways between sexes and between PCP, NIC and PCPNIC groups indicate functional overlaps. Distinctively, proteins differentially expressed exclusively in PCPNIC mice reveal unique effects associated with the comorbidity model. Interactome maps of these proteins identified sex-selective subnetworks, within which some proteins stood out: for females, peptidyl-prolyl cis-trans isomerase (Fkbp1a) and heat shock 70 kDa protein 1B (Hspa1b), both components of the oxidative stress subnetwork, and gamma-enolase (Eno2), a component of the energy metabolism subnetwork; and for males, amphiphysin (Amph), a component of the synaptic transmission subnetwork. These are proposed to be further investigated and validated as markers of the combined insult during adolescence.
Assuntos
Fenciclidina , Esquizofrenia , Camundongos , Animais , Masculino , Feminino , Fenciclidina/metabolismo , Esquizofrenia/metabolismo , Nicotina/farmacologia , Córtex Pré-Frontal/metabolismo , Transmissão Sináptica , Modelos Animais de DoençasRESUMO
The class of diterpenoids with a 14-carbon cembrane ring, the cembranoids, includes both competitive and noncompetitive inhibitors of the nicotinic acetylcholine receptor (AChR). All 20 coelenterate-derived cembranoids studied in this report inhibited [piperidyl-3,4-3H]-phencyclidine ([3H]-PCP) binding to its high-affinity site on the electric organ AChR, with IC50s ranging from 0.9 microM for methylpseudoplexaurate to 372 microM for lophotoxin. Inhibition was complete with all cembranoids but lophotoxin and most Hill coefficients were close to 1. Methylpseudoplexaurate and [3H]-PCP binding was competitive. Methylpseudoplexaurate and the fourth most potent cembranoid, eunicin, competed with each other for [3H]-PCP displacement, indicating that there exist one or more cembranoid sites on the AChR. Cembranoid affinity for the AChR correlated with hydrophobicity, but was also dependent on other features. Methylpseudoplexaurate and n-octanol also competed with each other for [3H]-PCP displacement, indicating that the cembranoid site is linked to the n-octanol site on the AChR. Unlike lophotoxin, the five cembranoids tested did not inhibit [125I]Tyr54-alpha-bungarotoxin binding to the AChR agonist sites. All seven cembranoids tested on oocyte-expressed electric organ AChR reversibly blocked acetylcholine-induced currents, although the inhibitor concentration curves were shallow and the inhibition was incomplete.
Assuntos
Diterpenos/farmacologia , Receptores Nicotínicos/efeitos dos fármacos , Acetilcolina/farmacologia , Animais , Sítios de Ligação , Bungarotoxinas/metabolismo , Feminino , Fenciclidina/metabolismo , Receptores Nicotínicos/metabolismo , Torpedo , Xenopus laevisRESUMO
This paper displays an attempt to elucidate the inhibitory mechanism of the nicotinic acetylcholine receptor (AChR) by spectroscopic means. Specifically, quantitative fluorescence spectroscopy was used to characterize: (1) the mechanism of quinacrine binding to its high-affinity noncompetitive inhibitor site located at the lipid-protein interface of the AChR and (2) the process by which agonists at high concentrations sterically compete for the quinacrine locus. For the first purpose, we study the temperature and ionic strength dependence of quinacrine binding by measuring the apparent dissociation constant (Kd) of quinacrine at the temperature range of 4-23 degrees C and in the sodium chloride (NaCl) concentration order of 0-250 mM. For the second objective, AChR native membranes from Torpedo californica electric organ suspended in buffer 10 mM sodium phosphate, pH 7.4, were preincubated with quinacrine for 2 h in the presence or in the absence of phencyclidine (PCP). Then, the PCP-sensitive quinacrine fluorescence was monitored while high concentrations of cholinergic agonists such as suberyldicholine, acetylcholine (ACh), or carbamylcholine were added to the suspension. By repeating these agonist back titrations at 4, 9, and 15 degrees C in the absence of NaCl and at 4 degrees C in the presence of 100 mM NaCl, we determined the temperature and ionic strength dependence of agonist binding to the quinacrine domain. These experiments suggest that the binding of both quinacrine (measured in the temperature range from 15 to 23 degrees C) and agonists at high concentrations (measured in the temperature regime of 4-15 degrees C) are enthalpy-driven processes, albeit that quinacrine binding is exothermic and agonist binding is endothermic. One plausible model to explain our results is that the quinacrine molecule needs first to be sterically well oriented to further enter into its binding site located in a crevice at the lipid-protein interface, whereas agonist molecules do not. Additionally, a relatively minimal electrostatic component is present in the quinacrine locus. Interestingly, the agonist inhibition constant values determined at 4 degrees C in the presence of 100 mM NaCl showed an exact correlation (slope = 1.03) with the reported concentration values of agonist that inhibit 50% of the maximum 86Rb+ efflux from AChR native vesicles in a 10-s assay with 80-85% of the alpha-bungarotoxin AChR sites occupied at zero membrane potential [S. A. Forman, L. L. Firestone, and K. W. Miller (1987) Biochemistry 26, 2807-2814]. This interdependence strongly supports the existence of a structural relationship between the agonist self-inhibitory binding site and the quinacrine locus. Although there exist evidence indicating that the process of agonist self-inhibition is mediated by a steric blockage of the ion channel, the occurrence of an agonist self-inhibitory binding site not located in the lumen channel indicates an allosteric mechanism for ion channel inhibition.
Assuntos
Agonistas Nicotínicos/metabolismo , Quinacrina/metabolismo , Receptores Nicotínicos/metabolismo , Animais , Sítios de Ligação , Órgão Elétrico/metabolismo , Técnicas In Vitro , Cinética , Modelos Biológicos , Concentração Osmolar , Fenciclidina/metabolismo , Cloreto de Sódio , Temperatura , Termodinâmica , Torpedo/metabolismoRESUMO
En la patologia esquizofrénica, los modelos bioquímicos explicativos mas comunes son: El de la Dopamina y el de la Fenciclidina. El neurotransmisor Dopamina actúa sobre los diversos receptores específicos, y la Fenciclidina sobre los llamados receptores de aminoácidos excitatorios. Otra hipótesis, como la de la Glicina o la de las Poliaminas, también tienen que ver con la acción sobre receptores de aminoacidos excitatorios. Esta revisión estudia algunos eventos neuroquímicos y neuropatológicos relacionados con los modelos mencionados. Conclusion: La esquizofrenia no puede ser explicada por un modelo simple, y mas bien es el resultado de una compleja interacción de disfunciones en los sistemas de neurotransmisión.